Flavell, Steven W et al. (2013) International Worm Meeting "Serotonin and PDF are opposing neuromodulators that control a bistable foraging behavior in C. elegans."

Pokala, Navin, Albrecht, Dirk A, Macosko, Evan Z, Bargmann, Cornelia I, Larsch, Johannes, Flavell, Steven W

[International Worm Meeting, 2013]

Foraging animals have distinct exploration and exploitation behaviors that are organized into discrete, long-lasting behavioral states. Here we characterize a neuromodulatory circuit that generates such long-lasting roaming and dwelling states in Caenorhabditis elegans. We find that two opposing neuromodulators, serotonin and the neuropeptide pigment dispersing factor (PDF), each initiate and extend one behavioral state. Serotonin promotes dwelling states through the MOD-1 serotonin-gated chloride channel. The spontaneous activity of serotonergic neurons correlates with dwelling behavior, and optogenetic modulation of the critical MOD-1-expressing targets induces long-lasting dwelling states. PDF promotes roaming states through the Gas-coupled PDFR-1 receptor; optogenetic activation of cAMP production in PDFR-1-expressing cells induces long-lasting roaming states. The neurons that produce and respond to each neuromodulator form a distributed circuit orthogonal to the classical wiring diagram, with several essential neurons that express each molecule. The slow temporal dynamics of the neuromodulatory circuit supplement fast motor circuits to give rise to long-lasting behavioral states.

Ji, Ni et al. (2017) International Worm Meeting "Long-term imaging of circuit-wide neural dynamics during unconstrained locomotion."

Flavell, Steven, Ji, Ni

[International Worm Meeting, 2017]

Mapping neural activity across the entire brain in a freely behaving animal represents one of the most ambitious and exciting challenges in neuroscience. In an effort to attain such ideal datasets in C. elegans, recent studies have established key technologies to image large numbers of neurons across the brain of C. elegans during locomotion. To allow this exciting approach to be robustly and routinely applied to circuit interrogation in C. elegans, we tackle a number of open technical challenges. These include 1) removing mechanical constraints to the animal during imaging, 2) improving light efficiency to minimize phototoxicity and bleaching, 3) reducing motion artifacts by improving behavioral tracking, and 4) developing computationally efficient software to automate the segmentation of neurons. We will present progress towards these goals and demonstrate the application of circuit-wide imaging during unconstrained locomotion.

Schier AF (2013) Cell "Should I stay or should I go: neuromodulators of behavioral states."

Schier AF

[Cell, 2013]

Animals are often in discrete behavioral states, but it is unclear how one specific state is generated and opposes alternative states. Flavell etal. now identify molecular and neural components in C.elegans that are involved in the generation of dwelling and roaming states.

McLachlan, Ian et al. (2021) International Worm Meeting "Diverse sensory cues and internal state converge on AWA chemoreceptor expression to enhance sensitivity to food odors"

Flavell, Steven, Kramer, Talya, Dua, Malvika, McLachlan, Ian

[International Worm Meeting, 2021]

To behave appropriately in dynamic environments, animals must integrate external stimuli with their recent experience and internal state. For example, successful foraging necessitates prioritizing food-related cues when hungry. To investigate how sensory experience and internal state can lead to lasting changes in nervous system function, we examined how recent experience impacts neuronal gene expression in the context of feeding behaviors. First, we used a ribosome-tagging approach to profile whole-nervous system gene expression in fed versus fasted animals. This approach identified 203 neuronal genes that are upregulated by fasting. Among these genes, G-protein coupled chemoreceptors putatively expressed in the AWA sensory neuron were overrepresented. We generated CRISPR knock-in transcriptional reporters for several of these chemoreceptors and confirmed their fasting-dependent upregulation in AWA. Of these, str-44 showed the most robust change in expression, so we performed in depth studies of its function and regulation. Strikingly, str-44 expression is influenced by a broad range of internal and external cues: the fed/fasted state of the animal, the presence of bacterial odors, and the presence of pathogens and additional stressors. Feeding state-dependent str-44 expression depends on insulin and TGF-beta signaling, as it is disrupted in daf-16/FOXO and daf-7/TGF-beta mutants. Experiments with food odors and non-ingestible bacteria suggest that, independent of feeding state, volatile food odors also impact str-44 expression. Chemosensory cues that influence str-44 expression are detected by AWA itself as well as additional tax-4-expressing neurons. Osmotic stress, heat stress, and infection by pathogenic bacteria also suppress the increase in str-44 expression during fasting, indicating that str-44 expression represents an integration of states rather than strictly nutritional state. AWA calcium responses to bacterial food are potentiated in the fasted state relative to the fed state, and this potentiation is reduced by osmotic stress during fasting. Preliminary experiments suggest that str-44 expression may be sufficient to confer sensitivity to putative food odors previously found to be detected by AWA with as of yet unidentified receptors. Based on these results, we suggest that environmental conditions that promote food-seeking behavior converge to regulate chemoreceptor expression in AWA, which in turn controls the animal's sensitivity to bacterial food.

Trojanowski, Nicholas et al. (2015) International Worm Meeting "Distinct mechanisms underlie two types of Caenorhabditis elegans sleep."

Raizen, David, Trojanowski, Nicholas, Fang-Yen, Christopher, Nelson, Matthew, Flavell, Steven

[International Worm Meeting, 2015]

Electrophysiological recordings have enabled identification of physiologically distinct yet behaviorally similar states of mammalian sleep. In contrast, sleep states in non-mammalian animals have generally been described behaviorally, and therefore non-mammalian sleep is often identified as a homogenous state characterized by quiescence of feeding and locomotion, reduced responsiveness, and rapid reversibility. In C .elegans, sleep has been described under two conditions: developmentally timed sleep (DTS or lethargus) occurs during molting transitions, while stress-induced sleep (SIS) occurs in response to exposure to cellular stress during adulthood. Behaviorally, DTS and SIS appear identical, as during both states feeding and locomotion cease and arousal threshold is increased. We used optogenetic manipulations of neuronal and muscular activity, pharmacology, and genetic perturbations to uncover circuit and molecular mechanisms of DTS and SIS. We find that locomotion quiescence induced by DTS- and SIS-associated neuropeptides (NLP-22 and FLP-13, respectively) occurs via their action on the nervous system. However, in mutants with activated Gs signaling, overexpression of NLP-22 or FLP-13 causes distinct locomotion phenotypes, suggesting that their neuronal target(s) and/or molecular mechanisms differ. Feeding quiescence during DTS results from a loss of pharyngeal muscle excitability, whereas feeding quiescence during SIS results from a loss of excitability in the nervous system. Together these results indicate that, as in mammals, distinct types of sleep in C. elegans are subserved by different mechanisms. .

Run JQ et al. (1992) Worm Breeder's Gazette "Genes That May Control the Cellular Sensation of Directionalitywithin C. elegans."

Hung M-S, Way JC, Run JQ, Wang AY

[Worm Breeder's Gazette, 1992]

unc-73 (e936 )animals have well-documented defects in the direction, and for some neurons, the extent of axon outgrowth (see Steven et al. abstract, this issue, for refs.). In addition, this strain shows, at a low penetrance, several phenotypes that are reminiscent of lin-17 (n671 ):production of additional mec-3 -expressingcells, mispositioning of PVM between its normal position and the AVM region, and secondary vulval protrusions. These observations suggest that unc-73 might be involved in the general sensation by cells of direction within C. elegans, and reduction of its function could lead to defects in axon outgrowth, cell migration, and asymmetric cell division. This notion is supported by the identification of a region of homology between UNC-73 and yeast Cdc24 p(Steven et al., this issue), which encodes an apparent guanine nucleotide exchange factor, and which is thought to control the sense of cell polarity within the yeast cell (cdc24 mutants are defective in both knowing where to place the bud and where to extend when shmooing). We envision that unc-73 might be part of a signal transduction pathway that would communicate directional information from extracellular gradients or the matrix to the cell's interior. To identify other genes in this pathway, we are pursuing two approaches. First, we have isolated non-Unc suppressors of unc-73 (e936 ).Most of these appear to be intragenic, but two suppressors appear to be extragenic, dominant, and map somewhere on chromosome II. These suppressors have no striking phenotype on their own. For a second approach, we sought to clone homologues of genes that interact with CDC24 .CDC42 is a member of the rho family of ras-like G proteins, and is a probable target of CDC24 .From the alignment of the yeast protein with a functional human CDC42 homologue, we designed degenerate oligonucleotides and PCR-amplified a ~700bp fragment. The sequence of each end of this fragment indicates that we probably have indeed cloned a C. elegans homologue of CDC42 .[See Figure]

Harris TW et al. (2000) J Cell Biol "Mutations in synaptojanin disrupt synaptic vesicle recycling."

Harris TW, Hartweig EA, Jorgensen EM, Horvitz HR

[J Cell Biol, 2000]

Synaptojanin is a polyphosphoinositide phosphatase that is found at synapses and binds to proteins implicated in endocytosis. For these reasons, it has been proposed that synaptojanin is involved in the recycling of synaptic vesicles. Here, we demonstrate that the unc-26 gene encodes the Caenorhabditis elegans ortholog of synaptojanin. unc-26 mutants exhibit defects in vesicle trafficking in several tissues, but most defects are found at synaptic termini. Specifically, we observed defects in the budding of synaptic vesicles from the plasma membrane, in the uncoating of vesicles after fission, in the recovery of vesicles from endosomes, and in the tethering of vesicles to the cytoskeleton. Thus, these results confirm studies of the mouse synaptojanin 1 mutants, which exhibit defects in the uncoating of synaptic vesicles (Cremona, O., G. Di Paolo, M.R. Wenk, A. Luthi, W.T. Kim, K. Takei, L. Daniell, Y. Nemoto, S.B. Shears, R.A. Flavell, D.A. McCormick, and P. De Camilli. 1999. Cell. 99:179-188), and further demonstrate that synaptojanin facilitates multiple steps of synaptic vesicle recycling.

Shuang Hu et al. (2008) C.elegans Neuronal Development Meeting "Regulation of Locomotion by UNC-73 RhoGEF-2 Isoforms"

Robert Steven, Tony Pawson, Shuang Hu

[C.elegans Neuronal Development Meeting, 2008]

unc-73 is a complex gene that functions in multiple tissue types at different stages of C. elegans development. It is required for normal fertility, pharynx and vulval muscle function and for specific cell migrations. In the nervous system unc-73 also plays a role in axon guidance and neurotransmission (Steven et al, 1998; Steven et al, 2005). unc-73 encodes at least eight protein isoforms that are differentially expressed. These isoforms are thought to function primarily through the activity of their respective RhoGEF domains, which catalyze the activation of members of either the Rac or Rho GTPase subfamilies. Here we attempt to better define the role of the UNC-73 isoforms in neurotransmission. Isoform specific rescue experiments previously revealed that the RhoGEF-2 domain-containing UNC-73C1, C2/F and E isoforms act redundantly to regulate the speed of locomotion. Transgenic animals lacking these isoforms have a lethargic movement phenotype that can be rescued by the expression of any individual isoform from this group. By using different cell-specific or inducible promoters to drive the expression of UNC-73E in transgenic unc-73 mutant animals we have begun to identify the temporal and spatial requirements of UNC-73E function. We have found that UNC-73E acts within the nervous system to physiologically regulate locomotion. It was recently reported that UNC-73E is an effector of Galphaq (Williams et al, 2007), however, it is still not clear how the UNC-73 isoforms function with Rho to regulate neurotransmission. In our hands the lethargic unc-73 mutants are not aldicarb resistant, which suggests that the UNC-73 RhoGEF-2 isoforms do not have an obvious defect in acetylcholine neurotransmitter release. This is consistent with the observation that restricted expression of UNC-73E in the cholinergic motor neurons does not rescue the unc-73lethargic movement phenotype. Epistasis analysis has revealed that activating mutations in different components of the Galphas pathway rescue the unc-73 lethargic movement phenotype caused by the loss of the UNC-73C1, C2/F and E isoforms. We are examining the possibility that these UNC-73 isoforms are required for the production or release of a neuromodulator that functions upstream of a Galphas-coupled receptor.

Antonopoulos CG (2016) Chaos "Dynamic range in the C. elegans brain network."

Antonopoulos CG

[Chaos, 2016]

We study external electrical perturbations and their responses in the brain dynamic network of the Caenorhabditis elegans soil worm, given by the connectome of its large somatic nervous system. Our analysis is inspired by a realistic experiment where one stimulates externally specific parts of the brain and studies the persistent neural activity triggered in other cortical regions. In this work, we perturb groups of neurons that form communities, identified by the walktrap community detection method, by trains of stereotypical electrical Poissonian impulses and study the propagation of neural activity to other communities by measuring the corresponding dynamic ranges and Steven law exponents. We show that when one perturbs specific communities, keeping the rest unperturbed, the external stimulations are able to propagate to some of them but not to all. There are also perturbations that do not trigger any response. We found that this depends on the initially perturbed community. Finally, we relate our findings for the former cases with low neural synchronization, self-criticality, and large information flow capacity, and interpret them as the ability of the brain network to respond to external perturbations when it works at criticality and its information flow capacity becomes maximal.

John Farver et al. (2007) International Worm Meeting "Specific UNC-73/Trio RhoGEF-2 isoforms function upstream of the Gas signaling pathway in neurons to physiologically regulate locomotion."

Tony Pawson, John Farver, Robert M. Steven

[International Worm Meeting, 2007]

unc-73 is a complex gene that functions in multiple tissue types at different stages of C. elegans development. unc-73 is essential for normal fertility, required for pharynx and vulval muscle function and for specific cell migrations and it plays a role in axon guidance and neurotransmission in the nervous system (Steven et al, 1998; Steven et al, 2005). unc-73 encodes at least eight protein isoforms that are differentially expressed. These isoforms are thought to function primarily through the activity of their respective RhoGEF domains, which catalyze the activation of members of either the Rac or Rho GTPase subfamilies. Here we attempt to better define the role of the UNC-73 isoforms in neurotransmission. Isoform specific rescue experiments previously revealed that the UNC-73C1, C2/F and E isoforms act redundantly to regulate the speed of locomotion. Transgenic animals lacking these isoforms have a lethargic movement phenotype that can be rescued by the expression of any individual isoform from this group. By using different cell-specific or inducible promoters to drive the expression of UNC-73E in transgenic unc-73 mutant animals we have started to narrow down the temporal and spatial requirements of UNC-73E function. We have found that UNC-73E acts within the nervous system, although not exclusively in the cholinergic motor neurons, to physiologically regulate locomotion. Epistasis analysis with other genes involved in the regulation of locomotion revealed that activating mutations in different components of the G<font face=symbol>a</font>_s pathway rescue the unc-73 lethargic movement phenotype caused by the loss of the UNC-73C1, C2/F and E isoforms. These data further point to a role for these isoforms in the regulation of synaptic activity. Since the G<font face=symbol>a</font>_s pathway acts primarily within the cholinergic motorneurons (Charlie et al, 2006), distinct from the site of activity of the UNC-73 isoforms, we propose that these isoforms function in neuromodulatory neurons, perhaps regulating neuromodulators or their receptors within these neurons. Along with the RhoGEF domains, unc-73 encodes other protein-protein interaction motifs including spectrin-repeat-like, SH3, Ig and FnIII domains. Using the yeast two-hybrid system we are in the process of screening for proteins that interact with the UNC-73 isoforms. The homologs of two of the proteins that we identified in our first screen are reported to function in vesicle trafficking, which may explain why unc-73 is involved in many cellular processes.